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A Rapid PCR‐based Method for the Identification of ob Mutant Mice
Author(s) -
Ellett Justin D.,
Evans Zachary P.,
Zhang Guojing,
Chavin Kenneth D.,
Spyropoulos Demetri D.
Publication year - 2009
Publication title -
obesity
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.438
H-Index - 199
eISSN - 1930-739X
pISSN - 1930-7381
DOI - 10.1038/oby.2008.443
Subject(s) - primer (cosmetics) , genotype , polymerase chain reaction , mutant , biology , allele , genetics , microbiology and biotechnology , gene , chemistry , organic chemistry
With increasing incidence of obesity, there is greater demand for suitable research and therapeutic models. The ob/ob mouse model develops obesity by 5 weeks of age. Previously, a method using DNA purification, PCR, and restriction digestion of products was devised to identify mice bearing the ob allele. Here, we describe a direct PCR method that requires no DNA purification. Wild‐type and ob‐specific primers are used under the same conditions in two separate and simultaneously run three‐primer PCRs. Standard PCR using the wild‐type primer mix produces 191 bp and 104 bp bands in +/+ and ob/+ and only the control 191 bp band in ob/ob animals. The ob‐specific primer reaction produces 191 bp and 123 bp bands in ob/+ and ob/ob and only the control 191 bp band in +/+ animals. Phenotypic weight gain in offspring of heterozygous intercrosses was used to validate genotypes. This primer‐specific PCR method allows simultaneous identification of +/+, ob/+, and ob/ob genotypes prior to breeding age to facilitate breeding and research studies in an important model of clinical obesity.